The primary objective of this project is to field test a molecular screening assay developed in our laboratory for the early detection of malignant and premalignant disease of the upper aerodigestive tract. A secondary goal is to identify molecular markers indicative of a high risk of tumor progression for premalignant lesions. These goals will be approached using three specific aims. In aim one, the molecular test using microsatellite alterations and tumor-specific hypermethylation to detect neoplastic cells exfoliated from the mucosa of at-risk individuals participating in community based cancer screening programs will be applied and evaluated. New molecular markers developed in Project 2 will be added to a panel of markers already validated in pilot studies involving individuals with known head and neck invasive cancer. The rate at which these alterations are present in the at-risk but presumably healthy population in the screening sessions will be determined together with the association of genetic alteration with exposure to known risk factors for head and neck cancer (HNSC). These healthy subjects will then be followed for 2 years and then reexamined. Specific aim #2 seeks to identify molecular markers for progression that will be combined together with physical findings and exposure history into a multivariate predictive model to investigate the independence of the molecular factors as predictors of disease progression. Individuals who are identified at screening sessions or in a consortium of medical and dental clinics to have visible oral premalignant or benign mucosal lesions will be eligible for participation in Specific aim #3 which will investigate molecular markers in the premalignant lesions and exfoliated cells again looking for markers associated with progression to invasive cancer. Molecular markers will be compared with histologic grade of the index lesion as well as risk-exposure history in multivariate analysis to identify the most useful predictors of progression. All molecular markers will be analyzed in extracted DNA taken from tissue biopsy, oral rinses and swabs, and peripheral blood plasma and compared with control DNA from peripheral blood leukocytes. Extracted DNA will be amplified using primers specific for microsatellites known to be frequently altered in HNSC and samples then separated on gels in order to identify tumorspecific alteration. Methylation markers will be assayed using Taqman fluorescence methodology. These studies promise to improve and validate an early detection paradigm that can then be applied in mass screening programs. The early detection of individuals likely to harbor HNSC through community screening programs would permit the diagnosis and treatment of the disease while it is still highly curable using current treatment methods. Accurate identification of individuals most likely to demonstrate progressive disease after presentation with premalignant lesions will permit the rational utilization of current and new treatment and preventive measures.